Numerous reports now implicate a loss of specialized cells termed interstitial cells of Cajal (ICC) in a variety of pediatric and adult gastrointestinal motor disorders. These disorders range from chronic intestinal pseudo- obstruction and meconium obstruction in infants, to gastroparesis both idiopathic and diabetic, megaduodenum, slow transit constipation and megacolon in adults. Current treatments of Gl complications have considerable side-effects and a greater understanding of the disease would provide novel targets for treatment and possible cure. Little or nothing is known about the fate of ICC following injury and whether ICC maintain the capacity to repopulate Gl tissues. The reported loss in Kit and disruption in ICC networks in humans and animal models exposes an imbalance in the normal homeodynamics of ICC in Gl tissues, which is likely a consequence of the failure to replace injured or dying ICC in Gl muscles. Loss of ICC has been attributed to cell death though the apoptotic signaling pathway or transdifferentiation. Using a multifaceted approach i.e. genetic, morphological and molecular approaches, including high throughput genetic screening and single cell genetic analysis, we addressed several fundamental questions that need answered. These questions included (i) What genes are preferentially expressed in ICC during their emergence from progenitors into adulthood that define their specialized phenotypes? (ii) What is the fate of ICC following their disruption and loss in the Gl tract? (iii) What are the factors that allow ICC to repopulate Gl muscles following their loss? Having developed powerful animal models, strains with cell-specific fluorescent reporters, expertise in enteric neurophysiology, interstitial cells, mechanisms of pacemaker function and neuroeffector transmission to better understand the diverse functional role of these cells in the Gl tract. Coupled with a full range of technologies to extensively evaluate ICC function including, electrophysiological techniques, morphological analysis, and dynamic Ca imaging. We believe it will take this degree of technological power to unravel the plasticity of ICC within the Gl tract. In summary, the completion of these specific aims will provide substantial insights into the underlying mechanisms of ICC development through to their mature phenotype. It will also provide important information on the cellular processes involved during ICC loss from Gl muscles and the exciting potential for recovery and re-emergence of ICC populations in Gl muscles. RELEVANCE (See instructions): Relevance to gastrointestinal motility disorders associated with a loss of interstitial cells of Cajal.